Identification of an unusual AT(D)Pase-like activity in multifunctional NAD glycohydrolase from the venom of Agkistrodon acutus.
NAD-glycohydrolases (NADases) are ubiquitous enzymes that possess NAD glycohydrolase, ADPR cyclase or cADPR hydrolase activity. All these activities are attributed to the NADase-catalyzed cleavage of C-N glycosyl bond. AA-NADase purified from the venom of Agkistrodon acutus is different from the known NADases, for it consists of two chains linked with disulfide-bond(s) and contains one Cu(2+) ion. Here, we show that AA-NADase is not only able to cleave the C-N glycosyl bond of NAD to produce ADPR and nicotinamide, but also able to cleave the phosphoanhydride linkages of ATP, ADP and AMP-PNP to yield AMP. AA-NADase selectively cleaves the P-O-P bond of ATP, ADP and AMP-PNP without the cleavage of P-O-P bond of NAD. The hydrolysis reactions of NAD, ATP and ADP catalyzed by AA-NADase are mutually competitive. ATP is the excellent substrate for AA-NADase with the highest specificity constant k(cat)/K(m) of 293+/-7mM(-1)s(-1). AA-NADase catalyzes the hydrolysis of ATP to produce AMP with an intermediate ADP. AA-NADase binds with one AMP with high affinity determined by isothermal titration calorimetry (ITC). AMP is an efficient inhibitor against NAD. AA-NADase has so far been identified as the first unique multicatalytic enzyme with both NADase and AT(D)Pase-like activities.
Zhang
Liyun
L
Department of Chemistry, University of Science and Technology of China, Hefei, Anhui, PR China.
Xu
Xiaolong
X
Luo
Zhaofeng
Z
Shen
Dengke
D
Wu
Hao
H
D036541:ADP-ribosyl Cyclase / Q000096:biosynthesis; D000244:Adenosine Diphosphate / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000378:metabolism; D017836:Agkistrodon / Q000378:metabolism*; D000818:Animals; D002384:Catalysis; D003300:Copper / Q000737:chemistry; D003435:Crotalid Venoms / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry; D006868:Hydrolysis; D009243:NAD / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D009244:NAD+ Nucleosidase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity
D003435:Crotalid Venoms; D004220:Disulfides; D009243:NAD; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D003300:Copper; D000255:Adenosine Triphosphate; D036541:ADP-ribosyl Cyclase; D009244:NAD+ Nucleosidase